title: NMO sera down-regulate AQP4 in human astrocyte and induce cytotoxicity independent of complement.
abstract: Autoantibodies against astrocyte water channel aquaporin-4 (AQP4) are highly specific for neuromyelitis optica (NMO). However, the molecular mechanism of NMO still remains unclear. The purpose of this study was to identify the possible humoral mechanisms responsible for the occurrence of astrocytic damage. Human primary astrocytes (AST) were immortalized by retroviral vectors harboring temperature-sensitive SV40 T antigen gene and AQP4 cDNA (M23), designated as hAST-AQP4. The effects of NMO sera on the content and localization of AQP4, including cytotoxicity and astrocytic morphology, were evaluated. In addition, this study examined whether the amount and localization of AQP4 protein in astrocytes were influenced by direct contact with the immortalized human brain microvascular endothelial cell line, TY09. NMO sera alone induced cytotoxicity and addition of complement had a more harmful effect on hAST-AQP4. NMO sera also decreased AQP4 mRNA and protein. NMO sera alone up-regulated TNFα and IL-6 in astrocytes and co-incubation with anti-TNFα and anti-IL-6 neutralizing antibodies blocked both the cytotoxicity and reduction of AQP4 in astrocytes. In the experiment using the in vitro BBB models, AQP4 protein mainly localized at the astrocytic membrane after co-culture with TY09, in contact with TY09. The future elucidation of factors that up-regulate AQP4 in astrocytes presumably released by blood brain barrier forming endothelial cells and that block the production of inflammatory cytokines may therefore lead to the development of a novel therapeutic strategy.
authors.affiliation: Department of Neurology and Clinical Neuroscience, Yamaguchi University Graduate School of Medicine, Ube, Yamaguchi, Japan.
authors.forename: Hiroyo
authors.initials: H
authors.lastname: Haruki
authors.forename: Yasuteru
authors.initials: Y
authors.lastname: Sano
authors.forename: Fumitaka
authors.initials: F
authors.lastname: Shimizu
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Omoto
authors.forename: Ayako
authors.initials: A
authors.lastname: Tasaki
authors.forename: Mariko
authors.initials: M
authors.lastname: Oishi
authors.forename: Michiaki
authors.initials: M
authors.lastname: Koga
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Saito
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Takahashi
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Nakada
authors.forename: Takashi
authors.initials: T
authors.lastname: Kanda
mesh_terms: D000906:Antibodies / Q000494:pharmacology; D051401:Aquaporin 4 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D001253:Astrocytes / Q000737:chemistry / Q000187:drug effects*; D002232:Carbocyanines / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D018920:Coculture Techniques; D003165:Complement System Proteins / Q000494:pharmacology*; D015536:Down-Regulation / Q000187:drug effects*; D042783:Endothelial Cells / Q000187:drug effects / Q000502:physiology; D004730:Endothelium, Vascular / Q000166:cytology; D027341:Excitatory Amino Acid Transporter 1 / Q000378:metabolism; D005904:Glial Fibrillary Acidic Protein; D006801:Humans; D015850:Interleukin-6 / Q000276:immunology; D009471:Neuromyelitis Optica / Q000097:blood*; D044967:Serum / Q000187:drug effects / Q000378:metabolism; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology
chemical_list: C496030:AQP4 protein, human; D000906:Antibodies; D051401:Aquaporin 4; D002232:Carbocyanines; D027341:Excitatory Amino Acid Transporter 1; D005904:Glial Fibrillary Acidic Protein; D015850:Interleukin-6; D014409:Tumor Necrosis Factor-alpha; C072024:carbocyanine dye DiIC12(3); D003165:Complement System Proteins
keywords: Aquaporin 4; Astrocyte; Blood–brain-barrier; Co-culture; Cytokine; Endothelial cell; Neuromyelitis optica